Synaptotagmin 1, a protein found on synaptic vesicles, serves as a Ca2+ sensor in neurotransmitter release. This protein has 16 homologues, and while these other proteins are widely thought to serve related functions essentially nothing is known about where they are or what they do. Preliminary data presented in this application shows that several synaptotagmin isoforms are abundant in the posterior and/or anterior pituitary, glands that release peptide hormones that control essentially all vital aspects of mammalian physiology. Prior to these new preliminary results, the pituitary had been established as a preparation amenable to quantitative biophysical study of how peptide-containing vesicles fuse with the plasma membrane to release their content into the circulation. Thus, the finding of novel synaptotagmin isoforms in the pituitary offers a unique opportunity to test their function in detail and elucidate their precise roles in excitation-secretion coupling. This project will study the pituitary synaptotagmin isoforms with complementary approaches, investigating their binding properties in vitro, and their localization and physiological function in the pituitary. The functional studies employ a strategy of using transgenic mice for gene ablation of specific synaptotagmin isoforms, and gene targeting to identify specific cell populations that express those isoforms for electrical recording. By performing capacitance recording in identified cells and nerve terminals we will determine how syt isoforms regulate exocytosis in terms of Ca2+ sensitivity and fusion pores. The experiments proposed here will test the functions of synaptotagmins 4, 7, 9, 10, 11, and 12 in the release of hormones that control growth, metabolism, stress, fluid balance, and reproduction. PUBLIC HEALTH RELEVANCE: These experiments will shed light on the function of a class of proteins with broad roles in neurological, mental, and endocrine function and thus contribute to improving treatment of neurological disorders and mental illness. By illuminating the mechanisms of control of the pituitary hormones oxytocin, vasopressin, growth hormone, adrenocorticotropin, thyroid stimulating hormone, follicle stimulating hormone, luteinizing hormone, and prolactin, these studies will improve our understanding of growth defects, abnormal stress responses, metabolic disorders, and reproductive health.